Center for Infectious Diseases, SUNY Stony Brook, CMM235, Stony Brook, NY 11794-5222, USA.
Infect Immun. 2011 Nov;79(11):4322-31. doi: 10.1128/IAI.05396-11. Epub 2011 Aug 15.
Yersinia pseudotuberculosis is a Gram-negative bacterial pathogen. Virulence in Y. pseudotuberculosis requires the plasmid-encoded Ysc type III secretion system (T3SS), which functions to translocate a set of effectors called Yops into infected host cells. The effectors function to antagonize phagocytosis (e.g., YopH) or to induce apoptosis (YopJ) in macrophages infected with Y. pseudotuberculosis. Additionally, when antiphagocytosis is incomplete and Y. pseudotuberculosis is internalized by macrophages, the bacterium can survive in phagosomes. Previous studies have shown that delivery of effectors into host cells occurs efficiently when Yersinia is extracellular. However, it is not clear whether the T3SS can be utilized by intracellular Y. pseudotuberculosis to translocate Yops. This possibility was investigated here using Y. pseudotuberculosis strains that express YopJ or YopH under the control of an inducible promoter. Bone marrow-derived murine macrophages were infected with these strains under conditions that prevented the survival of extracellular bacteria. Effector translocation was detected by measuring apoptosis or the activities of Yop-β-lactamase fusion proteins. Results showed that macrophages underwent apoptosis when YopJ expression was induced prior to phagocytosis, confirming that delivery of this effector prior to or during uptake is sufficient to cause cell death. However, macrophages also underwent apoptosis when YopJ was ectopically expressed after phagocytosis; furthermore, expression of the translocator YopB from intracellular bacteria also resulted in increased cell death. Analysis by microscopy showed that translocation of ectopically expressed YopH- or YopJ-β-lactamase fusions could be correlated with the presence of viable Y. pseudotuberculosis in macrophages. Collectively, our results suggest that the Ysc T3SS of Y. pseudotuberculosis can function within macrophage phagosomes to translocate Yops into the host cytosol.
假性结核耶尔森菌是一种革兰氏阴性细菌病原体。在假性结核耶尔森菌中，毒力需要质粒编码的 Ysc 型 III 型分泌系统 (T3SS)，该系统的功能是将一组效应物称为 Yops 易位到受感染的宿主细胞中。这些效应物的作用是拮抗吞噬作用（例如，YopH）或诱导感染假性结核耶尔森菌的巨噬细胞凋亡（YopJ）。此外，当吞噬作用不完全并且假性结核耶尔森菌被巨噬细胞内化时，细菌可以在吞噬体中存活。先前的研究表明，当耶尔森氏菌在细胞外时，效应物有效地递送到宿主细胞中。然而，尚不清楚 T3SS 是否可被细胞内的假性结核耶尔森菌利用来易位 Yops。在这里，使用表达受诱导型启动子控制的 YopJ 或 YopH 的假性结核耶尔森菌菌株进行了研究。在阻止细胞外细菌存活的条件下，用这些菌株感染骨髓来源的鼠巨噬细胞。通过测量凋亡或 Yop-β-内酰胺酶融合蛋白的活性来检测效应物的易位。结果表明，在吞噬作用之前诱导 YopJ 表达时，巨噬细胞发生凋亡，证实在此之前或期间递呈该效应物足以导致细胞死亡。然而，当吞噬作用后异位表达 YopJ 时，巨噬细胞也发生凋亡；此外，来自细胞内细菌的转运体 YopB 的表达也导致细胞死亡增加。显微镜分析表明，异位表达的 YopH 或 YopJ-β-内酰胺酶融合蛋白的易位与巨噬细胞中存活的假性结核耶尔森菌的存在相关。总之，我们的结果表明，假性结核耶尔森菌的 Ysc T3SS 可以在巨噬细胞吞噬体中发挥作用，将 Yops 易位到宿主细胞质中。
